The invention relates to a process for treating tissues using viable or nonviable cells, preferably microorganisms. More particularly, the invention relates to the use of microorganisms as an in vitro source of compositions for the processing of tissue to produce biomaterials useful in the production of bioprostheses.
Bioprostheses, i.e., bioprosthetic devices, are used to repair or replace damaged or diseased organs, tissues and other structures in humans and animals. Bioprostheses must be generally biocompatible with the recipient patient since they are typically implanted for extended periods of time. Bioprostheses can include artificial hearts, artificial heart valves, ligament repair material, vessel repair material, surgical patches constructed of mammalian tissue and the like. Considerable effort has been invested in the development of bioprosthetic heart valves, and the following discussion, for simplicity, will focus on these devices. It is to be understood, however, that the subject matter of this application is not limited to heart valves.
Currently available prostheses for the replacement of defective heart valves and other vascular structures may be classified as mechanical or bioprosthetic. Although mechanical valves have the advantage of proven durability through decades of use, they frequently are associated with a high incidence of blood clotting on or around the valve, necessitating continuous treatment with anticoagulants. Bioprosthetic heart valves constructed from materials derived from biological tissues were introduced in the early 1960's. Bioprosthetic heart valves are typically derived from porcine aortic valves or are manufactured from other biological materials such as bovine pericardium. Bioprostheses can include a combination of tissue-derived materials and synthetic materials.
A major rationale for the use of biological material for heart valves is that the profile and surface characteristics of biological material are optimal for laminar, nonturbulent flow. The result is that intravascular clotting is less likely to occur than with mechanical valves. This reduction in thrombogenicity has been well documented in clinical use of glutaraldehyde-fixed bioprosthetic valves. Glutaraldehyde fixes tissue by reacting to form covalent bonds with free amino groups in proteins, thereby chemically crosslinking nearby proteins.
Generally, bioprosthetic heart valves begin failing after about seven years following implantation, and few bioprosthetic valves remain functional after 20 years. Replacement of a degenerating valve prosthesis subjects the patient to additional surgical risk, especially in the elderly and in situations of emergency replacement. While failure of bioprostheses is a problem for patients of all ages, it is particularly pronounced in younger patients. Over fifty percent of bioprosthetic valve implants in patients under the age of 15 fail in less than five years, due to calcification.
Mineralization, e.g. calcification, appears to be the primary process leading to degeneration of bioprostheses. Efforts to address the calcification problem have included treating glutaraldehyde-fixed valves with compounds such as toluidine blue, sodium dodecyl sulfate and diphosphonate to reduce calcium nucleation. Other approaches include removal of reactive glutaraldehyde moieties from the tissue by a chemical process. Xenograft tissue, i.e., tissue from a species other than the species of the recipient patient, typically is fixed with glutaraldehyde prior to implantation to reduce the possibility of immunological rejection.
Still other approaches include development of alternative fixation techniques, since evidence suggests that the glutaraldehyde fixation process itself may contribute to calcification and mechanical deterioration. In addition, since nonviable cells present in transplanted tissue are sites for calcium deposition, investigators have developed various processes ("decellularization" processes) to remove cellular structure from the valve matrix. For example, detergents and nucleases have been used to obtain an extracellular matrix from tissue for use as graft material.
Fixation and treatment to reduce calcification generally involve harsh conditions that tend to sterilize the tissue. Absence of living microorganisms, primarily bacteria and fungi, is an important consideration for any bioprosthetic material intended for implantation into a patient since implantation of a nonsterile implant may be catastrophic for the patient. Thus, previous approaches to preparation of medical implants, including bioprostheses, generally have avoided introduction of microorganisms or, at least, have not encouraged the growth of microorganisms around and within the implant material.